期刊
ACS SUSTAINABLE CHEMISTRY & ENGINEERING
卷 8, 期 26, 页码 9763-9774出版社
AMER CHEMICAL SOC
DOI: 10.1021/acssuschemeng.0c02107
关键词
Poly-gamma-glutamic acid; Bacillus amyloliquefaciens; CRISPR-Cas9n; Jerusalem artichoke biorefinery
资金
- National Natural Science Foundation of China [21878152, 21776133]
- National Key Research and Development Program of China [2019YFA09005203]
- Natural Science Foundation of Jiangsu Province [BK20171010]
- Jiangsu Synergetic Innovation Center for Advanced Bio-Manufacture [XTB1804]
Bacillus amyloliquefaciens NB, a glutamate-independent poly-gamma-glutamic acid (gamma-PGA)-producing strain, can directly utilize inulin-containing sustainable materials. However, low gamma-PGA yield and lack of efficient genetic engineering approaches have hindered the industrial use of this strain. Here, we used the CRISPR-Cas9n technique to engineer B. amyloliquefaciens to enhance gamma-PGA production. We engineered three modules involved in inulin hydrolysis, reducing sugars metabolism, and gamma-PGA synthesis in B. amyloliquefaciens. Specifically, overexpresed the native inulin hydrolase CscA and two expression-optimized levanase and en doi nulinase, overexpressed of key genes related to reducing sugar metabolism to increased ATP production, and removed polysaccharide operon epsA-O and gamma-PGA hydrolase cwlO. Finally, the highest production of gamma-PGA (32.14 +/- 0.38 g/L) was obtained in a 7.5 L fed-batch fermenter. Thus, we successfully constructed an ideal candidate strain for efficient gamma-PGA production from inulin, which provides an important research basis for the development of more biobased products.
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